Several copper complexes have been assessed as anti-tumor agents against cancer cells. In this work, a copper compound [Cu(H2O){OS(CH3)2}L](NO3)2 incorporating the ligand 4′-phenyl-terpyridine antiproliferative activity against human colorectal, hepatocellular carcinomas and breast adenocarcinoma cell lines was determined, demonstrating high cytotoxicity. The compound is able to induce apoptosis and a slight delay in cancer cell cycle progression, probably by its interaction with DNA and induction of double-strand pDNA cleavage, which is enhanced by oxidative mechanisms. Moreover, proteomic studies indicate that the compound induces alterations in proteins involved in cytoskeleton maintenance, cell cycle progression and apoptosis, corroborating its antiproliferative potential.

Here we describe the establishment of a new canine mammary tumour (CMT) cell line, FR37-CMT that does not show dependence on female hormonal signaling to induce tumour xenografts in NOD-SCID mice. FR37-CMT cell line has a stellate or fusiform shape, displays the ability to reorganize the collagen matrix, expresses vimentin, CD44 and shows the loss of E-cadherin which is considered a fundamental event in epithelial to mesenchymal transition (EMT). The up-regulation of ZEB1, the detection of phosphorylated ERK1/2 and the downregulation of DICER1 and miR-200c are also in accordance with the mesenchymal characteristics of FR37-CMT cell line. FR37-CMT shows a higher resistance to cisplatin (IC50>50 µM) and to doxorubicin (IC50>5.3 µM) compared with other CMT cell lines. These results support the use of FR37-CMT as a new CMT model that may assist the understanding of the molecular mechanisms underlying EMT, CMT drug resistance, fostering the development of novel therapies targeting CMT.

Identification of novel molecules that can selectively inhibit the growth of tumor cells, avoid causing side effects to patients and/or intrinsic or acquired resistance, usually associated with common chemotherapeutic agents, is of utmost importance. Organometallic compounds have gained importance in oncologic chemotherapy, such as organotin(IV) complexes. In this study, we assessed the anti-tumor activity of the cyclic trinuclear organotin(IV) complex with an aromatic oximehydroxamic acid group [nBu2Sn(L)]3(H2L = N,2-dihydroxy-5-[N-hydroxyethanimidoyl]benzamide) – MG85 – and provided further characterization of its biological targets. We have previously shown the high anti-proliferative activity of this complex against human colorectal and hepatocellular carcinoma cell lines and lower cytotoxicity in neonatal non-tumor fibroblasts. MG85 induces tumor cell apoptosis and down-regulation of proteins related to tubulin dynamics (TCTP and COF1). Further characterization included the: (i) evaluation of interference in the cell cycle progression, including the expression of critical genes; (ii) affinity to DNA and the corresponding mode of binding; (iii) genotoxic potential in cells with deficient DNA repair pathways; and (iv) in vivo tumor reduction efficiency using mouse colorectal carcinoma xenografts.

The pH-sensitive affinity pair composed by neutravidin and iminobiotin was used to develop a multilayered Magnetic Resonance Imaging (MRI) nanoprobe responsive to the acidic pH of tumor microenvironment. The multilayer system was assembled on meso-2,3-dimercaptosuccinic acid-coated iron oxide magnetic nanoparticles (MNP), which convey negative MRI contrast enhancement properties to the nanoprobe. The outer stealth PEG-layer is altered in acidic media due to the disruption of interactions between neutravidin-iminobiotin. As a consequence, the positively charged inner layer is exposed and enhances interactions with cells. The nanoprobe uptake by HCT116 cells cultured in vitro under acidic conditions had a 2-fold increase compared to the uptake at physiological pH. The uptake difference is particularly clear in T2-weighted MRI phantoms of cells incubated with the nanoprobes at both pH conditions. This work sets the proof-of-concept of a MNP-based MRI nanoprobe targeting acidic tumor microenvironment through the use of a specific bio-recognition interaction that is pH-sensitive. This tumor targeting strategy is potentially applicable to the generality of tumors since the typical hypoxic conditions and high glycolysis rate in cancer cells create an acidic environment common to the majority of cancer types.

A series of Cu(diimine)(X-sal)(NO3) complexes, where the diimine is either 2,2'-bipyridine (bpy) or 1,10-phenanthroline (phen) and X-sal is a monoanionic halogenated salicylaldehyde (X = Cl, Br, I, or H), have been synthesized and characterized by elemental analysis and X-ray crystallography. Penta-coordinate geometries copper(II) were observed for all cases. The influence of the diimine coligands and different halogen atoms on the antiproliferative activities toward human cancer cell lines have been investigated. All Cu(II) complexes were able to induce a loss of A2780 ovarian carcinoma cell viability, with phen derivatives more active than bpy derivatives. In contrast, no in vitro antiproliferative effects were observed against the HCT116 colorectal cancer cell line. These cytotoxicity differences were not due to a different intracellular concentration of the complexes determined by inductively coupled plasma atomic emission spectroscopy. A small effect of different halogen substituents on the phenolic ring was observed, with X = Cl being the most highly active toward A2780 cells among the phen derivatives, while X = Br presented the lowest IC50 in A2780 cells for bpy analogs. Importantly, no reduction in normal primary fibroblasts cell viability was observed in the presence of bpy derivatives (IC50 > 40 muM). Mechanistically, complex 1 seems to induce a stronger apoptotic response with a higher increase in mitochondrial membrane depolarization and an increased level of intracellular reactive oxygen species (ROS) compared to complex 3. Together, these data and the low IC50 compared to cisplatin in A2780 ovarian carcinoma cell line demonstrate the potential of these bpy derivatives for further in vivo studies.

The syntheses of the heterometallic sodium and potassium-dioxidovanadium 2D polymers, [NaVO2(1kappaNOO';2kappaO"-L)(H2O)]n(1) and [KVO2(1kappaNOO';2kappaO';3kappaO"-L)(EtOH)]n(2) (where the kappa notation indicates the coordinating atoms of the polydentate ligand L) derived from (3,5-di-tert-butyl-2-hydroxybenzylidene)-2-hydroxybenzohydrazide (H2L) are reported. The polymers were characterized by IR, NMR, elemental analysis and single crystal X-ray diffraction analysis. The antiproliferative potential of 1 and 2 was examined towards four human cancer cell lines (ovarian carcinoma, A2780, colorectal carcinoma, HCT116, prostate carcinoma, PC3 and breast adenocarcinoma, MCF-7cell lines) and normal human fibroblasts. Complex 1 and 2 showed the highest cytotoxic activity against A2780 cell line (IC50 8.2 and 11.3muM, respectively) with 1>2 and an IC50 in the same range as cisplatin (IC50 3.4muM; obtained in the same experimental conditions) but, interestingly, with no cytotoxicity to healthy human fibroblasts for concentrations up to 75muM. This high cytotoxicity of 1 in ovarian cancer cells and its low cytotoxicity in healthy cells demonstrates its potential for further biological studies. Our results suggest that both complexes induce ovarian carcinoma cell death via apoptosis and autophagy, but autophagy is the main biological cause of the reduction of viability observed and that ROS (reactive oxygen species) may play an important role in triggering cell death.

Streptococcus dysgalactiae subsp. dysgalactiae (SDSD), a Lancefield group C streptococci (GCS), is a frequent cause of bovine mastitis. This highly prevalent disease is the costliest in dairy industry. Adherence and biofilm production are important factors in streptoccocal pathogenesis. We have previously described the adhesion and internalization of SDSD isolates in human cells and now we describe the biofilm production capability of this bacterium. In this work we integrated microbiology, imaging and computational methods to evaluate the biofilm production capability of SDSD isolates; to assess the presence of biofilm regulatory protein BrpA homolog in the biofilm producers; and to predict a structural model of BrpA-like protein and its binding to putative inhibitors. Our results show that SDSD isolates form biofilms on abiotic surface such as glass (hydrophilic) and polystyrene (hydrophobic), with the strongest biofilm formation observed in glass. This ability was mainly associated with a proteinaceous extracellular matrix, confirmed by the dispersion of the biofilms after proteinase K and trypsin treatment. The biofilm formation in SDSD isolates was also confirmed by confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Under SEM observation, VSD16 isolate formed cell aggregates during biofilm growth while VSD9 and VSD10 formed smooth and filmy layers. We show that brpA-like gene is present and expressed in SDSD biofilm-producing isolates and its expression levels correlated with the biofilm production capability, being more expressed in the late exponential phase of planktonic growth compared to biofilm growth. Fisetin, a known biofilm inhibitor and a putative BrpA binding molecule, dramatically inhibited biofilm formation by the SDSD isolates but did not affect planktonic growth, at the tested concentrations. Homology modeling was used to predict the 3D structure of BrpA-like protein. Using high throughput virtual screening and molecular docking, we selected five ligand molecules with strong binding affinity to the hydrophobic cleft of the protein, making them potential inhibitor candidates of the SDSD BrpA-like protein. These results warrant further investigations for developing novel strategies for SDSD anti-biofilm therapy.

Gold nanoparticles functionalized with thiolated oligonucleotides (Au-nanoprobes) have been used in a range of applications for the detection of bioanalytes of interest, from ions to proteins and DNA targets. These detection strategies are based on the unique optical properties of gold nanoparticles, in particular, the intense color that is subject to modulation by modification of the medium dieletric. Au-nanoprobes have been applied for the detection and characterization of specific DNA sequences of interest, namely pathogens and disease biomarkers. Nevertheless, despite its relevance, only a few reports exist on the detection of RNA targets. Among these strategies, the colorimetric detection of DNA has been proven to work for several different targets in controlled samples but demonstration in real clinical bioanalysis has been elusive. Here, we used a colorimetric method based on Au-nanoprobes for the direct detection of the e14a2 BCR-ABL fusion transcript in myeloid leukemia patient samples without the need for retro-transcription. Au-nanoprobes directly assessed total RNA from 38 clinical samples, and results were validated against reverse transcription-nested polymerase chain reaction (RT-nested PCR) and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The colorimetric Au-nanoprobe assay is a simple yet reliable strategy to scrutinize myeloid leukemia patients at diagnosis and evaluate progression, with obvious advantages in terms of time and cost, particularly in low- to medium-income countries where molecular screening is not routinely feasible.

MicroRNAs (miRNAs) regulate a wide variety of biological processes, including tumourigenesis. Altered miRNA expression is associated with deregulation of signalling pathways, which in turn cause abnormal cell growth and de-differentiation, contributing to cancer. miR-143 and miR-145 are anti-tumourigenic and influence the sensitivity of tumour cells to chemotherapy and targeted therapy. Comparative proteomic analysis was performed in HCT116 human colon cancer cells stably transduced with miR-143 or miR-145. Immunoblotting analysis validated the proteomic data in stable and transient miRNA overexpression conditions in human colon cancer cells. We show that approximately 100 proteins are differentially expressed in HCT116 human colon cancer cells stably transduced with miR-143 or miR-145 compared to Empty control cells. Further, Gene Ontology and pathway enrichment analysis indicated that proteins involved in specific cell signalling pathways such as cell death, response to oxidative stress, and protein folding might be modulated by these miRNAs. In particular, antioxidant enzyme superoxide dismutase 1 (SOD1) was downregulated by stable expression of either miR-143 or miR-145. Further, SOD1 gain-of-function experiments rescued cells from miR-143-induced oxidative stress. Moreover, miR-143 overexpression increased oxaliplatin-induced apoptosis associated with reactive oxygen species generation, which was abrogated by genetic and pharmacological inhibition of oxidative stress. Overall, miR-143 might circumvent resistance of colon cancer cells to oxaliplatin via increased oxidative stress in HCT116 human colon cancer cells.

Six new copper(ii) complexes with 2,2[prime or minute]:6[prime or minute],2[prime or minute][prime or minute]-terpyridine (4[prime or minute]-Rn-terpy) [1 (R1 = furan-2-yl), 2 (R2 = thiophen-2-yl), and 3 (R3 = 1-methyl-1H-pyrrol-2-yl)] and 2,6-di(thiazol-2-yl)pyridine derivatives (Rn-dtpy) [4 (R1), 5 (R2), and 6 (R3)] have been synthesized by a reaction between copper(ii) chloride and the corresponding ligand. The complexes have been characterized by UV-vis and IR spectroscopy, and their structures have been determined by X-ray analysis. The antiproliferative potential of copper(ii) complexes of 2,2[prime or minute]:6[prime or minute],2[prime or minute][prime or minute]-terpyridine and 2,6-di(thiazol-2-yl)pyridine derivatives towards human colorectal (HCT116) and ovarian (A2780) carcinoma as well as towards lung (A549) and breast adenocarcinoma (MCF7) cell lines was examined. Complex 1 and complex 6 were found to have the highest antiproliferative effect on A2780 ovarian carcinoma cells, particularly when compared with complex 2, 3 with no antiproliferative effect. The order of cytotoxicity in this cell line is 6 > 1 > 5 > 4 > 2 [approximate] 3. Complex 2 seems to be much more specific towards colorectal carcinoma HCT116 and lung adenocarcinoma A549 cells. The viability loss induced by the complexes agrees with Hoechst 33258 staining and typical morphological apoptotic characteristics like chromatin condensation and nuclear fragmentation. The specificity towards different types of cell lines and the low cytotoxic activity towards healthy cells are of particular interest and are a positive feature for further developments. Complexes 1-6 were also tested in the oxidation of alkanes and alcohols with hydrogen peroxide and tert-butyl-hydroperoxide (TBHP). The most active catalyst 4 gave, after 120 min, 0.105 M of cyclohexanol + cyclohexanone after reduction with PPh3. This concentration corresponds to a yield of 23% and TON = 210. Oxidation of cis-1,2-dimethylcyclohexane with m-CPBA catalyzed by 4 in the presence of HNO3 gave a product of a stereoselective reaction (trans/cis = 0.47). Oxidation of secondary alcohols afforded the target ketones in yields up to 98% and TON = 630.

A series of 2,2':6',2''-terpyridine (terpy), 2,6-di(thiazol-2-yl)pyridine (dtpy) and 2,6-di(pyrazin-2-yl)pyridine (dppy) derivatives with n-quinolyl substituents (n = 2 and 4) was used to synthesize five-coordinate complexes [CuCl2(n-quinolyl-terpy)] (1-2), [CuCl2(n-quinolyl-dtpy)] (3-4) and [CuCl2(n-quinolyl-dppy)] (5-6), respectively. The main emphasis of the research was to investigate the impact of the triimine skeleton (terpy, dtpy and dppy) and n-quinolyl pendant substituent on the antiproliferative and catalytic properties of 1-6. The obtained Cu(ii) compounds were studied as antiproliferative agents against human colorectal (HCT116) and ovarian (A2780) carcinoma, and they were used as catalysts for the oxidation of alkanes and alcohols with peroxides under mild conditions. The kinetic characteristics of the oxidizing species generated by the catalytic system Cu(ii) complex-H2O2 in CH3CN were obtained from the dependence of the alkane oxidation rate on its initial concentration. A model of competitive interaction of hydroxyl radicals with CH3CN and RH in the catalyst cavity has been proposed which is based on the simultaneous study of kinetics and selectivity in alkane oxidations.

Purpose: Progression of chronic myeloid leukemia (CML) is frequently associated with increased angiogenesis at the bone marrow mediated by exosomes. The capability of gold nanoparticles (AuNPs) functionalized with antiangiogenic peptides to hinder the formation of new blood vessels has been demonstrated in a chorioallantoic membrane (CAM) model. Methods: Exosomes of K562 CML cell line were isolated and their angiogenic effect assessed in a CAM model. AuNPs functionalized with antiangiogenic peptides were used to block the angiogenic effect of CML-derived exosomes, assessed by evaluation of expression levels of key modulators involved in angiogenic pathways - VEGFA, VEGFR1 (also known as FLT1) and IL8. Results: Exosomes isolated from K562 cells promoted the doubling of newly formed vessels associated with the increase of VEGFR1 expression. This is a concentration and time-dependent effect. The AuNPs functionalized with antiangiogenic peptides were capable to block the angiogenic effect by modulating VEGFR1 associated pathway. Conclusion: Exosomes derived from blast cells are capable to trigger (neo)-angiogenesis, a key factor for the progression and spreading of cancer, in particular in CML. AuNPs functionalized with specific antiangiogenic peptides are capable to block the effect of the exosomes produced by malignant cells via modulation of the intrinsic VEGFR pathway. Together, these data highlight the potential of nanomedicine-based strategies against cancer proliferation.

Gum arabic (GA) is a hydrophilic composite polysaccharide derived from exudates of Acacia senegal and Acacia seyal trees. It is biocompatible, possesses emulsifying and stabilizing properties and has been explored as coating agent of nanomaterials for biomedical applications, namely magnetic nanoparticles (MNPs). Previous studies focused on the adsorption of GA onto MNPs produced by co-precipitation methods. In this work, MNPs produced by a thermal decomposition method, known to produce uniform particles with better crystalline properties, were used for the covalent coupling of GA through its free amine groups, which increases the stability of the coating layer. The MNPs were produced by thermal decomposition of Fe(acac)3 in organic solvent and, after ligand-exchange with meso-2,3-dimercaptosuccinic acid (DMSA), GA coating was achieved by the establishment of a covalent bond between DMSA and GA moieties. Clusters of several magnetic cores entrapped in a shell of GA were obtained, with good colloidal stability and promising magnetic relaxation properties (r2 /r1 ratio of 350). HCT116 colorectal carcinoma cell line was used for in vitro cytotoxicity evaluation and cell-labeling efficiency studies. We show that, upon administration at the respective IC50 , GA coating enhances MNP cellular uptake by 19 times compared to particles bearing only DMSA moieties. Accordingly, in vitro MR images of cells incubated with increasing concentrations of GA-coated MNP present dose-dependent contrast enhancement. The obtained results suggest that the GA magnetic nanosystem could be used as a MRI contrast agent for cell-labeling applications.

Ruthenium-based drugs exhibit interesting properties as potential anticancer pharmaceuticals. We herein present the synthesis and characterization of a new family of ruthenium complexes with formulas [{Ru(bipy)2}2(μ-L)][CF3SO3]4 (L = bptz, 1a) and [{Ru(bipy)2}2(μ-L)][CF3SO3]2 (L = arphos, 2a; dppb, 3a; dppf, 4a), which were synthesized from the Ru(II) precursor compound cis-Ru(bipy)2Cl2. The complexes were characterized by elemental analysis, mass spectrometry, 1H and 31P{1H} NMR, IR spectroscopy, and conductivity measurements. The molecular structures for three Ru(II) compounds were determined by single-crystal X-ray diffraction. The newly developed compounds interact with CT-DNA by intercalation, in particular, 2a, 3a, and 4a, which also seemed to induce some extent of DNA degradation. This effect seemed to be related with the formation of reactive oxygen species. The cytotoxic activity was evaluated against A2780, MCF7, and MDAMB231 human tumor cells. Compounds 2a and 4a were the most cytotoxic with activity compared to cisplatin (∼2 μM, 72 h) in the A2780 cisplatin sensitive cells. All the compounds induced A2780 cell death by apoptosis, however, to a lesser extent for compounds 4a and 2a. For these compounds, the mechanism of cell death in addition to apoptosis seemed to involve autophagy. In vivo toxicity was evaluated using the zebrafish embryo model. LC50 estimates varied from 5.397 (3a) to 39.404 (1a) mg/L. Considering the in vivo toxicity in zebrafish embryos and the in vitro cytotoxicity in cancer cells, compound 1a seems to be the safest having no effect on dechirionation and presenting a good antiproliferative activity against ovarian carcinoma cells.Ruthenium-based drugs exhibit interesting properties as potential anticancer pharmaceuticals. We herein present the synthesis and characterization of a new family of ruthenium complexes with formulas [{Ru(bipy)2}2(μ-L)][CF3SO3]4 (L = bptz, 1a) and [{Ru(bipy)2}2(μ-L)][CF3SO3]2 (L = arphos, 2a; dppb, 3a; dppf, 4a), which were synthesized from the Ru(II) precursor compound cis-Ru(bipy)2Cl2. The complexes were characterized by elemental analysis, mass spectrometry, 1H and 31P{1H} NMR, IR spectroscopy, and conductivity measurements. The molecular structures for three Ru(II) compounds were determined by single-crystal X-ray diffraction. The newly developed compounds interact with CT-DNA by intercalation, in particular, 2a, 3a, and 4a, which also seemed to induce some extent of DNA degradation. This effect seemed to be related with the formation of reactive oxygen species. The cytotoxic activity was evaluated against A2780, MCF7, and MDAMB231 human tumor cells. Compounds 2a and 4a were the most cytotoxic with activity compared to cisplatin (∼2 μM, 72 h) in the A2780 cisplatin sensitive cells. All the compounds induced A2780 cell death by apoptosis, however, to a lesser extent for compounds 4a and 2a. For these compounds, the mechanism of cell death in addition to apoptosis seemed to involve autophagy. In vivo toxicity was evaluated using the zebrafish embryo model. LC50 estimates varied from 5.397 (3a) to 39.404 (1a) mg/L. Considering the in vivo toxicity in zebrafish embryos and the in vitro cytotoxicity in cancer cells, compound 1a seems to be the safest having no effect on dechirionation and presenting a good antiproliferative activity against ovarian carcinoma cells.

The major challenge of the pharmaceutical industry is to find potential solvents for poorly water-soluble drug molecules. Ionic liquids (ILs) have attracted this industry as (co-) solvents due to their unique physicochemical and biological properties. Herein, a straightforward approach for the enhancement of the water solubility of paracetamol and sodium diclofenac is presented, using new biocompatible N-acetyl amino acid N-alkyl cholinium-based ionic liquids as co-solvents (0.2-1mol%). These new ionic liquids were able to increase the water solubility of these drugs up to four times that in pure water or in an inorganic salt solution. In the presence of these ILs, the drugs lipophilicity (log P was not significantly changed for paracetamol, but for sodium diclofenac it was possible to decrease significantly its lipophilicity. Concerning cytotoxicity in human dermal fibroblasts it was observed that ILs did not show a significant toxicity, and were able to improve cell viability compared with the respective precursors.

Two mononuclear NiII and MnII compounds, [Ni(bdtbpza)2(CH3OH)4] (1) and [Mn(bdtbpza)2(CH3OH)2(H2O)2] (2), are afforded by employing a ‘scorpionate’ type precursor [bdtbpza = bis(3,5-di-t-butylpyrazol-1-yl)acetate]. The single crystal X-ray structure reveals that the central metal ion (NiII for 1 and MnII for 2) is surrounded by a pair of Oacetate atoms of two bis(pyrazol-1-yl)acetate units, while four OMeOH donors for 1 and two OMeOH plus two Owater for 2 complete the first coordination sphere. Thus, both compounds exhibit a slightly distorted octahedral geometry possessing an O6 coordination environment. EPR spectra of CuII-doped 1 and of 2 recorded on the polycrystalline solids and in organic solution confirm the octahedral geometry around the metal ions and the binding of six oxygen atoms. The electrochemical study of compounds 1 and 2 shows that one electron reduction of MnII occurs at a more negative potential than NiII, indicating a lower tendency of reduction for Mn than Ni. Both compounds displayed a high cytotoxic activity against A2780 ovarian carcinoma cells and no cytotoxic activity in normal primary human fibroblasts for concentrations up to 55 μM. Notwithstanding, compound 1 is found to be the most cytotoxic towards A2780 cancer cells. The cytotoxic activity of compound 1 is correlated with the induction of apoptosis associated with a higher mitochondria dysfunction and autophagy cell death. In addition, the compounds can induce oxidative damage leading to ROS accumulation. Overall, the data presented here demonstrate that 1 has potential for further in vivo studies aiming at its future application in ovarian cancer therapy.

Silver nanoparticles (AgNPs) were prepared by GREEN chemistry relying on the reduction of AgNO3 by phytochemicals present in black tea extract. AgNPs were fully characterized by transmission electron microscopy (TEM), ultraviolet-visible spectroscopy ((UV-vis)), X-ray diffraction (XRD) and energy dispersive absorption spectroscopy (EDS). The synthesized AgNPs induced a decrease of the cell viability in a dose-dependent manner with a low IC50 (0.5+/-0.1muM) for an ovarian carcinoma cell line (A2780) compared to primary human fibroblasts (IC50 5.0+/-0.1muM). The DNA binding capability of CT (calf thymus) DNA was investigated using electronic absorption and fluorescence spectroscopies, circular dichroism and viscosity titration methods. Additionally, the AgNPs strongly quench the intrinsic fluorescence of BSA, as determined by synchronous fluorescence spectra.